CN109392892B - Biological collagen-based material preserving fluid and application thereof - Google Patents
Biological collagen-based material preserving fluid and application thereof Download PDFInfo
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- CN109392892B CN109392892B CN201811535544.2A CN201811535544A CN109392892B CN 109392892 B CN109392892 B CN 109392892B CN 201811535544 A CN201811535544 A CN 201811535544A CN 109392892 B CN109392892 B CN 109392892B
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N1/00—Preservation of bodies of humans or animals, or parts thereof
- A01N1/02—Preservation of living parts
- A01N1/0205—Chemical aspects
- A01N1/021—Preservation or perfusion media, liquids, solids or gases used in the preservation of cells, tissue, organs or bodily fluids
- A01N1/0221—Freeze-process protecting agents, i.e. substances protecting cells from effects of the physical process, e.g. cryoprotectants, osmolarity regulators like oncotic agents
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N1/00—Preservation of bodies of humans or animals, or parts thereof
- A01N1/02—Preservation of living parts
- A01N1/0205—Chemical aspects
- A01N1/021—Preservation or perfusion media, liquids, solids or gases used in the preservation of cells, tissue, organs or bodily fluids
- A01N1/0226—Physiologically active agents, i.e. substances affecting physiological processes of cells and tissue to be preserved, e.g. anti-oxidants or nutrients
Abstract
The invention discloses a preservation solution for a biological extracellular matrix material, which consists of antifreeze, a structure protective agent, an anticoagulant, an antioxidant and EDTA-2Na, and can improve the low-temperature tolerance of a preserved object through the combined action of the antifreeze, the structure protective agent and the anticoagulant, limit the chain segment motion of the preserved object to keep the surface biological performance of the preserved object, and prevent oxidative deterioration in the irradiation process under the synergistic action of the antioxidant and the EDTA-2 Na. The preparation method is simple, low in cost and convenient to use, can reduce equipment and power consumption required by preservation, is nontoxic and micromolecules, can be absorbed and metabolized by organisms, reduces the time and difficulty required by pretreatment of preoperative samples, and is more convenient for clinical use.
Description
Technical Field
The invention belongs to the field of biomedical materials, and particularly relates to a biological collagen-based material preserving fluid and application thereof.
Background
The biological collagen-based material has important clinical value and is widely applied to regenerative medicine and tissue engineering, but the long-term storage of the biomembrane material in the biological collagen-based material is still a problem which is difficult to overcome. Soft bioactive materials such as cornea, amnion, xeno-mucosa, etc. must be preserved in a wet state in order to maintain morphology and activity, otherwise the surface may collapse and seriously affect the effect if water is lost. When the bioactive material is placed in the special preserving liquid during wet preservation, the preserving liquid can play a role in infiltration and isolation, uniform pressure can be applied to the surface of a sample, the sample can be fully stretched to expose active structure sites of the sample, the sample is simultaneously coagulated towards the inside by uniform external force, and the appearance and the function can be fully maintained.
However, the movement of the material in the preservation solution is not static, and the microstructure of the material can still move continuously and generate irreversible entanglement to gradually wrap the active sites of the material, so that the effectiveness of the material is influenced. To solve these problems, generally employed methods are to lower the temperature or to use a high viscosity preserving fluid, in both of which brownian motion of the living segment can be effectively restricted, thereby extending the active retention time of the material. The most commonly used preserving fluid is glycerin, the preserving time can be prolonged to more than one year when the temperature is reduced to below-40 ℃ in the process of temperature reduction and preservation of common glycerin preserving fluid, the preserving fluid can be maintained for more than three years at below-80 ℃, and the preserving fluid needs to be preserved in liquid nitrogen for a longer time. If the preserving fluid with certain viscosity is adopted, the chain motion and the interlacing of the material can be limited, the maximum temperature of half-year storage can be increased to 0 ℃, and the storage and transportation difficulty is reduced.
Sodium hyaluronate is the most commonly used material for preparing high viscosity preserving fluid, has the advantages of no toxicity and good affinity, and is also commonly used as a raw material of viscoelastic agent in ophthalmology. Sodium hyaluronate has high viscosity at low concentration, and can effectively limit chain motion and coagulation. However, biological products must be sterilized before being used as drugs or devices for clinical use for safety and long-term storage, and radiation sterilization is the only viable method. Various high molecular materials such as sodium hyaluronate and the like can be degraded under the irradiation effect, so that the viscosity is rapidly reduced to ensure that the preservation solution is ineffective, the degradation is uncontrollable, and an active intermediate generated in the process reacts with a biological sample in the preservation solution to influence the product quality. In animal experiments and clinical application, instability of part of products in use is probably caused by the influence of macromolecular preservative fluid. Researchers also find that the biological collagen-based material preserved by the macromolecular preservative solution needs a longer time to be treated and prepared before the operation, because the macromolecules attached to the surface of the material can be adsorbed and combined with the surface of the product under the action of irradiation, and tissues need a longer time to complete surface cleaning and regenerative repair. In addition, in the macromolecule preservation solution, because the macromolecule production method is generally fermentation or biological extraction, some glycoprotein, foreign protein and the like may be mixed in the material to cause allergy; the high viscosity also makes the colloidal liquid unevenly distributed, and air bubbles are easily mixed during subpackage to influence the quality after irradiation.
Therefore, the preservation solution capable of overcoming the problems is developed, so that the preservation and transportation cost can be reduced, and the quality stability of the product can be improved to ensure the operation quality and the treatment effect. However, at present, no other method capable of well replacing macromolecular preservation solution for long-term preservation of biological materials exists, and most of non-macromolecular preservation solutions disclosed by the patents are used for medium-term preservation, namely preservation at 2-8 ℃ for less than 1 month; the cornea storage liquid disclosed in CN201410344982 can be stored for about 1 year when the cornea storage liquid is matched with a high-viscosity storage liquid and is possibly kept below-20 ℃; CN201510995653 likewise achieves long-term maintenance of the placenta by means of a high viscosity preservation solution and low temperature. In the invention, the combination of the anti-agglomeration agent and the antioxidant with a certain concentration is found by the research of technical personnel, so that the biological collagen-based material can achieve the effect of limiting the chain segment movement, and the influence of irradiation on the stability of a sample can be reduced.
Disclosure of Invention
In order to solve the influence of the conventional macromolecular preservative solution on the biological collagen-based material, the invention aims to provide the special preservative solution for the biological collagen-based material, which is composed of a low-molecular-weight material.
The invention also aims to provide application of the biological collagen-based material preservation solution.
The technical scheme adopted by the invention is as follows:
a biological collagen-based material preservation solution is an aqueous solution composed of the following components in parts by mass: 100-700 g/L of antifreeze, 5-200 g/L of structure protective agent, 10-100 g/L of anticoagulant, 0.1-10 g/L, EDTA-2Na 0.1-5 g/L of antioxidant, and the pH value of aqueous solution is 6.5-7.5.
Preferably, the anti-freezing liquid is glycerol, propylene glycol or a mixture of glycerol and propylene glycol.
Preferably, the structure protective agent is a low molecular weight sugar or sugar alcohol.
Preferably, the number of glycosidically linked monomers in the low molecular weight sugar or sugar alcohol structure does not exceed 3. The low molecular weight sugars or sugar alcohols include, but are not limited to, glucose, fructose, mannose, sucrose, trehalose, maltose, raffinose, erythritol, xylitol, mannitol, sorbitol, maltitol.
Preferably, the anticoagulant is sodium citrate, sodium tartrate or sodium malate.
Preferably, the antioxidant comprises cysteine, N-acetylcysteine and sodium thiosulfate.
The biological collagen-based material preservation solution is applied to preservation of biological collagen-based materials.
Preferably, the collagen-based material comprises small intestine submucosa, amnion, biological cornea and pericardium, cartilage, nerve and blood vessel epimatrix.
The biological collagen-based material preservation solution is mainly used for low-temperature medium-long term preservation of materials such as small intestine submucosa, amnion, biological cornea and pericardium, cartilage, nerve, blood vessel epimatrix and the like.
Preferably, the low temperature is a temperature of not higher than 0 ℃.
In the invention, glycerin, propylene glycol or a mixture of glycerin and propylene glycol are used as main components of the preservation solution, so that the preservation solution can prevent freezing of a preserved sample. In the cryopreservation of biological materials, whether living tissue with cells or extracellular matrix with cells, the most significant effect on their structure and activity is ice crystallization, since the crystallization of water causes volume changes that severely damage the protected tissue structure. The freezing point of the pure glycerol is 20 ℃, and the freezing point of the aqueous solution of the pure glycerol is lower than-20 ℃ when the mass fraction of the aqueous solution is 45-80%, so the pure glycerol is widely used in low-temperature medium-term storage. However, since different segments of the protein may have different effects on glycerol, the preserved biological sample may locally adsorb too much glycerol or repel glycerol, and the glycerol is unevenly distributed in the preserved biological sample, so that the internal structure may be deformed over a long time to damage the sample structure. Therefore, after the collagen-based material is stored in pure glycerol and a pure glycerol-water system for a long time at a non-ultralow temperature, the collagen-based material can expand, stretch and break inside the system, so that the structure of a collagen network is damaged. The freezing point of the propylene glycol is-59 ℃, the anti-freezing effect is better than that of the glycerol, but the viscosity is low, the toughening effect is weaker, and the integral performance of the sample is influenced to a certain extent. In the present invention, the addition of low molecular weight sugars or sugar alcohols can improve this phenomenon. This is because low molecular sugar or sugar alcohol has moderate hydrogen bond strength and high affinity with collagen material, sugar/sugar alcohol in propylene glycol-sugar/sugar alcohol-water system can toughen collagen, while sugar/sugar alcohol in glycerin-sugar/sugar alcohol-water system can reduce adsorption between glycerin and collagen, and in propylene glycol-glycerin-sugar/sugar alcohol mixed system, both sugar/sugar alcohol and propylene glycol contribute to excessive adsorption of dispersed glycerin in collagen network, thereby improving collagen preservation performance. In the invention, low molecular weight sugar or sugar alcohol is selected as a structure protective agent, so that the surface stability of the biological extracellular matrix sample at low temperature can be effectively protected. It is generally considered that multiple layers of water molecules are distributed on the surface of a protein molecule, water molecules around the protein molecule are frozen continuously in the process of cooling, and if a single layer of water molecules on the surface of the protein molecule is also frozen, hydrogen bonds and polar groups on the surface of the protein are denatured. Glycerol can also be regarded as a three-carbon sugar alcohol, but the hydrogen bond forming capability of the glycerol is very strong, after a sample is placed in a glycerol solution, the surface water layer and the inner water layer can be quickly and completely replaced by the glycerol, and local enrichment can occur to cause regionalization, crystallization and damage to the structure at low temperature for a long time. The low molecular weight saccharides and sugar alcohols also have a large amount of hydroxyl groups, and the hydrogen bonding effect between the low molecular weight saccharides and sugar alcohols is weaker than that of glycerol, so that a hydration film can be formed after partial replacement of the water on the surface of protein, but the hydrogen bonding strength and density are weaker, and the regional crystallization cannot be sufficiently maintained at low temperature for a long time, so that the surface of the sample can be protected. The oligomers such as monosaccharide, disaccharide, trisaccharide and low molecular weight sugar alcohol have the structure of polyhydroxy aldehyde ketone, have moderate hydrogen bond strength and high affinity with collagen materials, and have a protective effect on protein possibly related to the chirality of low molecular weight sugar.
In the present invention, sodium citrate, sodium tartrate or sodium malate is preferably used as the anticoagulant. In aqueous solution, the surface chain segment of the collagen matrix material can be physically entangled due to Brownian motion, and the existence of polar groups on the chain or adsorption to form ions with charges can aggravate the physical entanglement, so that the bioactivity of the material is reduced. And the counter ions are distributed around the charged ions on the chain, the counter ions form an adsorption layer or a diffusion layer on the surface or around the surface of the counter ions, and the potential difference between the adsorption layer and the diffusion layer outside the adsorption layer and the diffusion layer is called Zeta potential. Higher concentrations of sodium citrate, sodium tartrate or sodium malate can raise the Zeta potential, i.e., stabilize the ions, resist aggregation and physical entanglement of the segments, and prevent surface changes from affecting material properties.
In the invention, the reason for adding the antioxidants of cysteine and sodium thiosulfate is that free radicals can be generated in the preservation solution due to the existence of dissolved oxygen and under the final irradiation action of the sample, and active groups such as amino and sulfhydryl are likely to be oxidized and denatured under the action of the free radicals, even chain scission and crosslinking can occur to influence the quality stability of the sample. Cysteine, N-acetylcysteine and sodium thiosulfate are common non-toxic antioxidants, can prevent a sample from being excessively oxidized after irradiation, and are non-toxic and do not influence clinical use.
In the invention, EDTA-2Na can play an auxiliary role in antioxidation. The strong complexation of EDTA can prevent the catalytic action of metal ions on the free radical chain reaction, thereby inhibiting the oxidation reaction. Similarly, sodium citrate, sodium tartrate or sodium malate may also undergo some complexation with metal ions to synergistically enhance the efficacy of the antioxidant.
In the invention, the preservation solution is generally used for preserving biological collagen-based materials without maintaining cell activity, such as biological cornea, amniotic membrane, nerve, cartilage materials and the like, the temperature is generally required to be kept below 0 ℃, and the preservation time can reach 0.5-1 year.
Compared with the existing materials, the invention has the beneficial effects that:
the invention provides a biological collagen-based material preservation solution, which can ensure that biological products such as biological cornea, amnion, nerve, bone extracellular matrix and the like have stable quality during preservation, the structure is well protected, the long-term preservation time below 0 ℃ can reach 6 months to 1 year, and the surface structure and activity can be well preserved.
The invention improves the defects of the prior common glycerin preserving fluid. The shape of the product can be changed when the common glycerol preservation solution is used for preserving the soft sample for too long, and the method can effectively limit the aggregation of sample chain segments and the surface denaturation caused by physical crosslinking, thereby prolonging the low-temperature preservation time of the product.
The invention avoids the quality problem possibly brought by the high-molecular high-viscosity preserving fluid. The viscosity of the preservation solution with the main component of the macromolecule can be sharply reduced under irradiation to generate a large amount of uncontrollable degradation products, the antioxidant is difficult to fully exert, and adverse effects on the quality can be caused by the reaction of the antioxidant on the collagen material. In addition, the common polymer materials are usually obtained by fermentation or animal extraction, and impurities in the common polymer materials also increase the quality risk of the product. The invention does not adopt high molecular materials, but adds high-concentration functional components, thereby not only preventing the surface performance and the internal structure of the sample from being damaged, but also preventing the irradiation oxidation from influencing the sample.
The invention has the beneficial effects that:
the components of the preservation solution are nontoxic micromolecules, so that the preservation solution is convenient for clinical application and reduces the pretreatment time of samples. The conventional high-viscosity preservation solution may require a relatively long time for sample pretreatment during clinical use, and the sample quality may even be seriously affected by the sample treatment process. The invention can be directly used for operation by taking out the preserved object and simply washing, is convenient to use and has few influence factors on quality. Meanwhile, the biopolymer material with higher price is not added, the price of each component is low, the preservation solution is simple to prepare and has wide applicability, and the preservation solution is very suitable for preserving various collagen-based biological products.
Drawings
FIG. 1 is a graph showing the effect of example 5, pure glycerin and pure propylene glycol, on biological corneas before and after 1 year of storage.
Detailed Description
The present invention will be further described with reference to specific examples, but the present invention is not limited thereto.
Example 1
Taking 700g of propylene glycol, 5g of glucose, 10g of sodium tartrate, 0.1g of cysteine and 0.1g of EDTA-2Na, adjusting the pH of the solution to 6.5-7.0, and diluting the solution to 1L by using a buffer solution with the same pH, wherein the buffer solution can be used for storing samples such as amniotic membrane, small intestine submucosa and the like.
Example 2
Taking 100g of glycerol, 200g of fructose, 100g of sodium citrate, 10g of sodium thiosulfate and 5.0g of EDTA-2Na, adjusting the pH of the solution to 7.0-7.5, and diluting the solution to 1L by using a buffer solution with the same pH for storing samples such as small intestine submucosa, pericardium epimatrix and the like.
Example 3
Taking 500g of glycerol, 10g of mannose, 20g of sodium malate, 5g of cysteine and 2.0g of EDTA-2Na, adjusting the pH of the solution to 6.5-7.0, and diluting the solution to 1L by using a buffer solution with the same pH for preserving samples such as biological cornea, amniotic membrane and the like.
Example 4
Taking 330g of glycerol, 120g of propylene glycol, 50g of sucrose, 15g of sodium malate, 4.5g of sodium thiosulfate and 2.0g of EDTA-2Na, adjusting the pH of the solution to 7.0-7.5, and diluting the solution to 1L by using a buffer solution with the same pH for preserving samples such as biological cornea, amniotic membrane, small intestine submucosa and the like.
Example 5
480g of glycerol, 55g of propylene glycol, 25g of trehalose, 20g of sodium tartrate, 7.5g of N-acetylcysteine and 3.5g of EDTA-2Na are taken, the pH of the solution is adjusted to 6.5-7.0, and then the solution is diluted to 1L by using a buffer solution with the same pH value and is used for preserving samples such as cartilage extracellular matrix, biological nerve, biological cornea and the like.
Example 6
170g of glycerol, 130g of propylene glycol, 120g of maltose, 60g of sodium citrate, 3.0g of sodium thiosulfate and 1.5g of EDTA-2Na are taken, the pH of the solution is adjusted to 7.0-7.5, and then the solution is diluted to 1L by a buffer solution with the same pH value and is used for preserving samples such as cartilage extracellular matrix, biological nerve, biological cornea and the like.
Example 7
90g of glycerol, 310g of propylene glycol, 40g of raffinose, 25g of sodium malate, 2.5g of cysteine and 1.2g of EDTA-2Na are taken, the pH of the solution is adjusted to 6.5-7.0, and then the solution is diluted to 1L by using a buffer solution with the same pH value and is used for preserving samples such as cartilage extracellular matrix, biological nerve, biological cornea and the like.
Example 8
Taking 50g of glycerol, 550g of propylene glycol, 25g of erythritol, 15g of sodium tartrate, 0.5g of sodium thiosulfate and 0.3g of EDTA-2Na, adjusting the pH of the solution to 7.0-7.5, and diluting the solution to 1L by using a buffer solution with the same pH, wherein the buffer solution is used for preserving samples such as cartilage extracellular matrix, biological nerve, biological cornea and the like.
Example 9
Taking 120g of glycerol, 230g of propylene glycol, 35g of xylitol, 70g of sodium citrate, 4.5g of N-acetylcysteine and 2.5g of EDTA-2Na, adjusting the pH of the solution to 6.5-7.0, and diluting the solution to 1L by using a buffer solution with the same pH value for preserving samples such as biological cornea, amniotic membrane and the like.
Example 10
Taking 250g of glycerol, 250g of propylene glycol, 85g of mannitol, 20g of sodium citrate, 6g of sodium thiosulfate and 3.0g of EDTA-2Na, adjusting the pH of the solution to 7.0-7.5, and diluting the solution to 1L by using a buffer solution with the same pH value for preserving samples such as small intestine submucosa, amnion, blood vessel epimatrix and the like.
Example 11
Taking 75g of glycerol, 75g of propylene glycol, 150g of sorbitol, 90g of sodium malate, 9g of cysteine and 4.5g of EDTA-2Na, adjusting the pH of the solution to 6.5-7.0, and diluting the solution to 1L by using a buffer solution with the same pH for preserving samples such as small intestine submucosa, amnion, blood vessel epimatrix and the like.
Example 12
The method comprises the steps of taking 100g of glycerol, 100g of propylene glycol, 120g of maltitol, 80g of sodium tartrate, 8g of sodium thiosulfate and 4.0g of EDTA-2Na, adjusting the pH of the solution to 7.0-7.5, and diluting the solution to 1L by using a buffer solution with the same pH, wherein the buffer solution is used for preserving samples such as biological cornea, small intestine submucosa, extracellular matrix and the like.
Testing
The preservation solutions described in examples 1 to 12 were prepared and used for preserving a biological collagen matrix material sample, and the biological cornea and amniotic membrane were preserved using common pure glycerol, pure propylene glycol, glycerol cryopreservation solution (glycerol-DMEM culture solution volume ratio 1: 1), propylene glycol preservation solution (example 1 with glucose removed), and high-viscosity preservation solution (containing hyaluronic acid concentration 15g/L, glycerol concentration 350g/L, and chondroitin sulfate 15 g/L) as control examples. Subpackaging the preserved materials in suitable containers, sterilizing by irradiation, and storing at 0 deg.C. The most intuitive change of the surface properties of the biological cornea, the small intestine submucosa and the amniotic membrane is whether yellowing occurs or not, and the surface of the sample gradually changes from colorless to brown after surface crosslinking, physical entanglement or oxidation, so three observation points of irradiation, low-temperature storage for half a year and low-temperature storage for one year are selected to observe the change condition of the sample. The experimental results are shown in table one.
The preservation effect of example 5, pure glycerol and pure propylene glycol on biological cornea is shown in fig. 1.
As can be seen from the data in the table, the biological samples can be stored for 6 months to 1 year at the temperature of below 0 ℃ in all the embodiments, the surface morphology of the biological samples has no obvious change, and the biological samples can be widely applied to the medical field. Examples 1, 4, 12 showed a slight yellowing at 1 year of storage, which may be related to the lack of viable cell components in fresh amniotic membrane, but no significant change in other forms. The test results of the high-viscosity preserving fluid are combined, so that the preserving effect of the low-molecular-weight preserving fluid can reach or be better than that of the high-viscosity preserving fluid, the effect of the high-viscosity preserving fluid is actually better than that of the low-molecular-weight preserving fluid when the high-molecular-weight preserving fluid is not irradiated, but the viscosity of the preserving fluid is sharply reduced after the high-molecular-weight preserving fluid is irradiated, and the property of viscosity preservation is lost.
In the comparative example, when the pure glycerol is used for storing samples, the water on the surfaces of the samples is replaced, so that the irradiation does not obviously affect the samples, and the surfaces of the samples are not yellowed after the irradiation is finished, but the water layers on the surfaces of the samples can be damaged too strongly due to the hydrogen bond action of the glycerol during long-time storage, so that the mechanical property and the surface property of the stored products are also affected. The glycerol frozen stock solution contains complex biological components, so that complex free radicals and autoxidation reaction can occur during irradiation, thereby seriously affecting the performance of the preserved object and being not suitable for the preservation of the extrabiological matrix material needing irradiation treatment. Indeed, pure glycerol and glycerol cryopreservation solutions may be more suitable for non-irradiation long-term low temperature cryopreservation of live cell-containing materials below-40 ℃, and the storage time above 0 ℃ will generally not exceed 1 month.
The pure propylene glycol of the comparative example has good antifreezing property, but has poor protective effect on samples, can not limit the free movement of collagen segments, and also has radiation resistance, so that the preserved objects are yellowed. In contrast, the propylene glycol preservation solution of the comparative example reduced glucose as a structure protectant as compared with example 1, and the color of the amnion was darker and the surface properties were affected after long-term storage, although the shape of the amnion was not significantly changed during long-term storage.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention are all equivalent substitutions which are included in the protection scope of the present invention.
Claims (3)
1. A biological collagen-based material preservation solution is an aqueous solution composed of the following components in parts by mass:
100-700 g/L of antifreeze, 5-200 g/L of structure protective agent, 10-100 g/L of anticoagulant, 0.1-10 g/L, EDTA-2Na 0.1-5 g/L of antioxidant, and the pH value of aqueous solution is 6.5-7.5; the antifreezing solution is glycerol, propylene glycol or a mixture of glycerol and propylene glycol; the structure protective agent is low molecular weight sugar or sugar alcohol; the number of monomers connected by glycosidic bonds in the low molecular weight sugar or sugar alcohol structure is not more than 3;
the anticoagulant is sodium citrate, sodium tartrate or sodium malate;
the antioxidant is cysteine, N-acetylcysteine and sodium thiosulfate.
2. The use of the preservation solution of a biocollagen-based material according to claim 1 in the preservation of a biocollagen-based material; the biological collagen-based material is small intestine submucosa, amnion, biological cornea and pericardium, cartilage, nerve and blood vessel epimatrix.
3. Use according to claim 2, characterized in that the preservation temperature is not higher than 0 ℃.
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| CN109939263A (en) * | 2019-03-29 | 2019-06-28 | 广州锐澄医疗技术有限公司 | A kind of preparation method of heterogeneity biological amnion product |
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Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101502257A (en) * | 2009-03-13 | 2009-08-12 | 厦门大学 | Corneal midterm preservation solution and preparation method thereof |
| CN101720753A (en) * | 2009-12-09 | 2010-06-09 | 中国人民解放军第四军医大学 | Cryopreservation solution of tissue engineering products and application method thereof |
| CN102132697A (en) * | 2010-01-21 | 2011-07-27 | 周海华 | Amniotic membrane long-term preserving fluid and preparation method thereof |
| CN102308787A (en) * | 2011-07-06 | 2012-01-11 | 中国人民解放军第四军医大学 | Cartilage preservation solution |
| CN104094925A (en) * | 2014-07-18 | 2014-10-15 | 广州优得清生物科技有限公司 | Lamellar corneal preserving solution |
| CN104938478A (en) * | 2015-05-20 | 2015-09-30 | 泰山医学院 | Cartilago articularis vitrification cryoprotectant and cartilage preservation method |
| CN104938476A (en) * | 2015-03-02 | 2015-09-30 | 泰山医学院 | Osteochondral graft preservation liquid and preparation method thereof |
-
2018
- 2018-12-14 CN CN201811535544.2A patent/CN109392892B/en active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101502257A (en) * | 2009-03-13 | 2009-08-12 | 厦门大学 | Corneal midterm preservation solution and preparation method thereof |
| CN101720753A (en) * | 2009-12-09 | 2010-06-09 | 中国人民解放军第四军医大学 | Cryopreservation solution of tissue engineering products and application method thereof |
| CN102132697A (en) * | 2010-01-21 | 2011-07-27 | 周海华 | Amniotic membrane long-term preserving fluid and preparation method thereof |
| CN102308787A (en) * | 2011-07-06 | 2012-01-11 | 中国人民解放军第四军医大学 | Cartilage preservation solution |
| CN104094925A (en) * | 2014-07-18 | 2014-10-15 | 广州优得清生物科技有限公司 | Lamellar corneal preserving solution |
| CN104938476A (en) * | 2015-03-02 | 2015-09-30 | 泰山医学院 | Osteochondral graft preservation liquid and preparation method thereof |
| CN104938478A (en) * | 2015-05-20 | 2015-09-30 | 泰山医学院 | Cartilago articularis vitrification cryoprotectant and cartilage preservation method |
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